Resident angiogenic mesenchymal stem cells from multiorgan donor thoracic aortas

Stem cells are one of the most fascinating areas of biology today, and since the discover of an adult population, i.e., adult Stem Cells (aSCs), they have generated much interest especially for their application potential as a source for cell based regenerative medicine and tissue engineering. aSCs have been found in different tissues including bone marrow, skin, intestine, central nervous system, where they reside in a special microenviroment termed “niche” which regulate the homeostasis and repair of adult tissues. The arterial wall of the blood vessels is much more plastic than ever before believed. Several animal studies have demonstrated the presence of cells with stem cell characteristics within the adult vessels. Recently, it has been also hypothesized the presence of a “vasculogenic zone” in human adult arteries in which a complete hierarchy of resident stem cells and progenitors could be niched during lifetime. Accordingly, it can be speculated that in that location resident mesenchymal stem cells (MSCs) with the ability to differentiate in smooth muscle cells, surrounding pericytes and fibroblasts are present. The present research was aimed at identifying in situ and isolating MSCs from thoracic aortas of young and healthy heart-beating multiorgan donors. Immunohistochemistry performed on fresh and frozen human thoracic aortas demonstrated the presence of the vasculogenic zone between the media and the adventitial layers in which a well preserved plexus of CD34 positive cells was found. These cells expressed intensely HLA-I antigens both before and after cryopreservation and after 4 days of organ cultures remained viable. Following these preliminary results, we succeeded to isolate mesenchymal cells from multi-organ thoracic aortas using a mechanical and enzymatic combined procedure. Cells had phenotypic characteristics of MSC i.e., CD44+, CD90+, CD105+, CD166+, CD34low, CD45- and revealed a transcript expression of stem cell markers, e.g., OCT4, c-kit, BCRP-1, IL6 and BMI-1. As previously documented using bone marrow derived MSCs, resident vascular wall MSCs were able to differentiate in vitro into endothelial cells in the presence of low-serum supplemented with VEGF-A (50 ng/ml) for 7 days. Under the condition described above, cultured cells showed an increased expression of KDR and eNOS, down-regulation of the CD133 transcript, vWF expression as documented by flow cytometry, immunofluorescence, qPCR and TEM. Moreover, matrigel assay revealed that VEGF induced cells were able to form capillary-like structures within 6 hours of seeding. In summary, these findings indicate that thoracic aortas from heart-beating, multi-organ donors are highly suitable for obtaining MSCs with the ability to differentiate in vitro into endothelial cells. Even though their differentiating potential remains to be fully established, it is believed that their angiogenic ability could be a useful property for allogenic use. These cells can be expanded rapidly, providing numbers which are adequate for therapeutic neovascularization; furthermore they can be cryostored in appropriate cell banking facilities for later use.

Trapianto di polmone ed ex-vivo. Esperienza iniziale in due centri trapianti di polmone

Background Il trapianto polmonare rappresenta l'unica soluzione percorribile per l'insufficienza respiratoria end-stage. La principale limitante ad un suo impiego è rappresentata dalla scarsità di polmoni idonei. In questo contesto si colloca il progetto di ricondizionamento ex-vivo. L'implementazione risulta possibile attraverso il ricondizionamento di organi cosiddetti “marginali” o giudicati inizialmente non idonei al trapianto e da donatori a cuore fermo (Donor after Cardiac Death – DCD). Scopo Lo scopo principale del progetto è quello di analizzare i risultati a breve e medio termine del trapianto polmonare effettuato impiegando polmoni sottoposti a procedura di ricondizionamento ex-vivo in due centri trapianti di polmone a basso volume. Materiali e Metodi I dati sono stati raccolti retrospettivamente (da Giugno 2013 per quanto riguarda l’ISMETT e dal 2014 quelli dell’ Ospedale S. Orsola-Malpighi) e in maniera prospettica dal 2019. Risultati Sommando i dati relativi ad entrambi i centri, dal 2013 sono stati eseguiti un totale di 26 procedure di riperfusione. Sei volte i polmoni sono stati giudicati idonei al trapianto al termine con 5 trapianti doppi e un trapianto singolo sinistro. I risultati in termine di sopravvivenza a medio termine sono risultati sovrapponibili a quelli disponibili in letteratura, tuttavia è stato registrato un alto tasso di PGD (Primary Graft Disfunction) di grado 3, da imputare alla patologia di base del ricevente (ipertensione polmonare), che per protocollo di istituto rimane con supporto cardio-respiratorio (ECMO) dopo il trapianto. Conclusioni L’EVLP si conferma in centri di piccolo e medio volume un metodo sicuro ed efficace per implementare in numero di polmoni idonei al trapianto.